Wednesday, November 30, 2016

Contact electrification of individual dielectric microparticles measured by optical tweezers in air

Haesung Park and Thomas W. LeBrun

We measure charging of single dielectric microparticles after interaction with a glass substrate using optical tweezers to control the particle, measure its charge with a sensitivity of a few electrons and precisely contact the particle with the substrate. Polystyrene (PS) microparticles adhered to the substrate can be selected based on size, shape, or optical properties and repeatedly loaded into the optical trap using a piezoelectric (PZT) transducer. Separation from the substrate leads to charge transfer through contact electrification. The charge on the trapped microparticles is measured from the response of the particle motion to a step excitation of a uniform electric field. The particle is then placed onto a target location of the substrate in a controlled manner. Thus, the triboelectric charging profile of the selected PS microparticle can be measured and controlled through repeated cycles of trap loading followed by charge measurement. Reversible optical trap loading and manipulation of the selected particle leads to new capabilities to study and control successive and small changes in surface interactions.


Optical manipulation using highly focused alternate radially and azimuthally polarized beams modulated by a devil’s lens

Zhirong Liu and P. H. Jones

We propose and demonstrate a novel high numerical aperture (NA) focusing system composed of an annular beam with alternate radially and azimuthally polarized rings, focused by a devil’s lens (DL), and further investigate its radiation forces acting upon a Rayleigh particle both analytically and numerically. Strongly focused cylindrical vector beams produce either dark-centered or peak-centered intensity distributions depending on the state of polarization, whereas the DL produces a series of foci along the propagation direction. We exploit these focusing properties and show that by selecting an appropriate truncation parameter in front of the focusing lens, the proposed optical focusing system can selectively trap and manipulate dielectric micro-particles with low or high refractive indices by simply adjusting the radius of the pupil or the beam. Finally, the stability conditions for effectively trapping and manipulating Rayleigh particles are analyzed. The results obtained in this work are of interest in possible applications in optical confinement and manipulation, sorting micro-particles, and making use of a DL.


Fast and Accurate Algorithm for Repeated Optical Trapping Simulations on Arbitrarily Shaped Particles Based on Boundary Element Method

Kai-Jiang Xu, Xiao-Min Pan, Ren-Xian Li, Xin-Qing Sheng

In optical trapping applications, the optical force should be investigated within a wide range of parameter space in terms of beam configuration to reach the desirable performance. A simple but reliable way of conducting the related investigation is to evaluate optical forces corresponding to all possible beam configurations. Although the optical force exerted on arbitrarily shaped particles can be well predicted by boundary element method (BEM), such investigation is time costing because it involves many repetitions of expensive computation, where the forces are calculated from the equivalent surface currents. An algorithm is proposed to alleviate the difficulty by exploiting our previously developed skeletonization framework. The proposed algorithm succeeds in reducing the number of repetitions. Since the number of skeleton beams is always much less than that of beams in question, the computation can be very efficient. The proposed algorithm is accurate because the skeletonization is accuracy controllable.


Tuesday, November 29, 2016

The RNA helicase Mtr4p is a duplex-sensing translocase

Eric M Patrick, Sukanya Srinivasan, Eckhard Jankowsky & Matthew J Comstock

The conserved Saccharomyces cerevisiae Ski2-like RNA helicase Mtr4p plays essential roles in eukaryotic nuclear RNA processing. RNA helicase activity of Mtr4p is critical for biological functions of the enzyme, but the molecular basis for RNA unwinding is not understood. Here, single-molecule high-resolution optical trapping measurements reveal that Mtr4p unwinds RNA duplexes by 3′-to-5′ translocation on the loading strand, that strand separation occurs in discrete steps of 6 base pairs and that a single Mtr4p molecule performs consecutive unwinding steps. We further show that RNA unwinding by Mtr4p requires interaction with upstream RNA duplex. Inclusion of Mtr4p within the TRAMP complex increases the rate constant for unwinding initiation but does not change the characteristics of Mtr4p's helicase mechanism. Our data indicate that Mtr4p utilizes a previously unknown unwinding mode that combines aspects of canonical translocating helicases and non-canonical duplex-sensing helicases, thereby restricting directional translocation to duplex regions.


Ripplon laser through stimulated emission mediated by water waves

Samuel Kaminski, Leopoldo L. Martin, Shai Maayani & Tal Carmon

Lasers rely on stimulated electronic transition, a quantum phenomenon in the form of population inversion. In contrast, phonon masers1, 2, 3 depend on stimulated Raman scattering and are entirely classical. Here we extend Raman lasers1, 2, 3 to rely on capillary waves, which are unique to the liquid phase of matter and relate to the attraction between intimate fluid particles. We fabricate resonators that co-host capillary4 and optical modes5, control them to operate at their non-resolved sideband and observe stimulated capillary scattering and the coherent excitation of capillary resonances at kilohertz rates (which can be heard in audio files recorded by us). By exchanging energy between electromagnetic and capillary waves, we bridge the interfacial tension phenomena at the liquid phase boundary to optics. This approach may impact optofluidics by allowing optical control, interrogation and cooling6 of water waves.


Lab on Fiber Technology for biological sensing applications

Patrizio Vaiano, Benito Carotenuto, Marco Pisco, Armando Ricciardi, Giuseppe Quero, Marco Consales, Alessio Crescitelli, Emanuela Esposito, Andrea Cusano

This review presents an overview of “Lab on Fiber” technologies and devices with special focus on the design and development of advanced fiber optic nanoprobes for biological applications. Depending on the specific location where functional materials at micro and nanoscale are integrated, “Lab on Fiber Technology” is classified into three main paradigms: Lab on Tip (where functional materials are integrated onto the optical fiber tip), Lab around Fiber (where functional materials are integrated on the outer surface of optical fibers), and Lab in Fiber (where functional materials are integrated within the holey structure of specialty optical fibers).
This work reviews the strategies, the main achievements and related devices developed in the “Lab on Fiber” roadmap, discussing perspectives and challenges that lie ahead, with special focus on biological sensing applications.


Multibuilding Block Janus Synthesized by Seed-Mediated Self-Assembly for Enhanced Photothermal Effects and Colored Brownian Motion in an Optical Trap

Kanokwan Sansanaphongpricha, Michael C. DeSantis, Hongwei Chen, Wei Cheng, Kai Sun, Bo Wen, Duxin Sun

The asymmetrical features and unique properties of multibuilding block Janus nanostructures (JNSs) provide superior functions for biomedical applications. However, their production process is very challenging. This problem has hampered the progress of JNS research and the exploration of their applications. In this study, an asymmetrical multibuilding block gold/iron oxide JNS has been generated to enhance photothermal effects and display colored Brownian motion in an optical trap. JNS is formed by seed-mediated self-assembly of nanoparticle-loaded thermocleavable micelles, where the hydrophobic backbones of the polymer are disrupted at high temperatures, resulting in secondary self-assembly and structural rearrangement. The JNS significantly enhances photothermal effects compared to their homogeneous counterpart after near-infrared (NIR) light irradiation. The asymmetrical distribution of gold and iron oxide within JNS also generates uneven thermophoretic force to display active colored Brownian rotational motion in a single-beam gradient optical trap. These properties indicate that the asymmetrical JNS could be employed as a strong photothermal therapy mediator and a fuel-free nanoscale Janus motor under NIR light.


Monday, November 28, 2016

One-step separation-free detection of carcinoembryonic antigen in whole serum: Combination of two-photon excitation fluorescence and optical trapping

Cheng-Yu Li, Di Cao, Chu-Bo Qi, Hong-Lei Chen, Ya-Tao Wan, Yi Lin, Zhi-Ling Zhang, Dai-Wen Pang, Hong-Wu Tang

Direct analysis of biomolecules in complex biological samples remains a major challenge for fluorescence-based approaches due to the interference of background signals. Herein, we report an analytical methodology by exploiting a single low-cost near-infrared sub-nanosecond pulse laser to synchronously actualize optical trapping and two-photon excitation fluorescence for senstive detection of carcinoembryonic antigen (CEA) in buffer solution and human whole serum with no separation steps. The assay is performed by simultaneously trapping and exciting the same immune-conjugated microsphere fabricated with a sandwich immunization strategy. Since the signal is strictly limited in the region of a three-dimensional focal volume where the microsphere is trapped, no obvious background signal is found to contribute the detected signals and thus high signal-to-background data are obtained. As a proof-of-concept study, the constructed platform exhibits good specificity for CEA and the detection limit reaches as low as 8 pg/mL (45 fM) with a wide linear range from 0.01 to 60 ng/mL in the both cases. To investigate the potential application of this platform in clinical diagnosis, 15 cases of serum samples were analyzed with satisfactory results, which further confirm the applicability of this method.


Computation of radiation pressure force exerted on arbitrary shaped homogeneous particles by high-order Bessel vortex beams using MLFMA

Minglin Yang, Yueqian Wu, Kuan Fang Ren, and Xinqing Sheng

Due to special characteristics of nondiffraction and self reconstruction, the Bessel beams have attracted wide attention in optical trapping and appear to be a dramatic alternative to Gaussian beams. We present in this paper an efficient approach based on the surface integral equations (SIE) to compute the radiation pressure force (RPF) exerted on arbitrary shaped homogeneous particles by high-order Bessel vortex beam (HOBVB). The incident beam is described by vector expressions perfectly satisfy Maxwell’s equations. The problem is formulated with the combined tangential formulation (CTF) and solved iteratively with the aid of the multilevel fast multipole algorithm (MLFMA). Then RPF is computed by vector flux of the Maxwell’s stress tensor over a spherical surface tightly enclosing the particle and analytical expression for electromagnetic fields of incident beam in near region are used. The numerical predictions are compared with the results of the rigorous method for spherical particle to validate the accuracy of the approach. Some numerical results on relative large particles of complex shape, such as biconcave cell-like particles with different geometry parameters are given, showing powerful capability of our approach. These results are expected to provide useful insights into the RPF exerted on complex shaped particles by HOBVB.


Strong light confinement and gradient force in a hexagonal boron nitride slot waveguide

Bofeng Zhu, Guobin Ren, Yixiao Gao, Haisu Li, Beilei Wu, and Shuisheng Jian

In this Letter, we show that a hexagonal boron nitride (h-BN) slot waveguide can achieve strong field enhancement and light confinement in a slot region and a giant gradient force between h-BN slabs. Excellent agreement between simulations and results from an analytical model is observed. In a two-dimensional case, a field enhancement ratio near 60, a power confinement ratio of 80%, and a gradient force over −8.5 nN/μm×mW−8.5 nN/μm×mW have been achieved, which are much higher than the slot waveguide based on artificial hyperbolic metamaterials. The gradient force and power confinement ratio in a three-dimensional slot waveguide structure are also studied. A gradient force of −1.2 nN/mW−1.2 nN/mW and a power confinement ratio of 50% have been obtained. The h-BN slot waveguide may have potential in particle manipulation.


Suppression of photothermal convection of microparticles in two dimensional nanoplasmonic optical lattice

Yi-Chung Chen, Gilad Yossifon and Ya-Tang Yang
Photothermal convection has been a major obstacle for stable particle trapping in plasmonic optical tweezer at high optical power. Here, we demonstrate a strategy to suppress the plasmonic photothermal convection by using vanishingly small thermal expansion coefficient of water at low temperature. A simple square nanoplasmonic array is illuminated with a loosely Gaussian beam to produce a two dimensional optical lattice for trapping of micro particles. We observe stable particle trapping due to near-field optical gradient forces at elevated optical power at low temperature. In contrast, for the same optical power at room temperature, the particles are convected away from the center of the optical lattice without their accumulation. This technique will greatly increase usable optical power and enhance the trapping capability of plasmonic optical tweezer.


Tuesday, November 22, 2016

Optical Torques on Upconverting Particles for Intracellular Microrheometry

Paloma Rodríguez-Sevilla, Yuhai Zhang, Nuno de Sousa, Manuel I. Marqués, Francisco Sanz-Rodríguez, Daniel Jaque, Xiaogang Liu, and Patricia Haro-González

Precise knowledge and control over the orientation of individual upconverting particles is extremely important for full exploiting their capabilities as multifunctional bioprobes for interdisciplinary applications. In this work, we report on how time-resolved, single particle polarized spectroscopy can be used to determine the orientation dynamics of a single upconverting particle when entering into an optical trap. Experimental results have unequivocally evidenced the existence of a unique stable configuration. Numerical simulations and simple numerical calculations have demonstrated that the dipole magnetic interactions between the upconverting particle and trapping radiation are the main mechanisms responsible of the optical torques that drive the upconverting particle to its stable orientation. Finally, how a proper analysis of the rotation dynamics of a single upconverting particle within an optical trap can provide valuable information about the properties of the medium in which it is suspended is demonstrated. A proof of concept is given in which the laser driven intracellular rotation of upconverting particles is used to successfully determine the intracellular dynamic viscosity by a passive and an active method.


Hidden Markov Modeling with Detailed Balance and Its Application to Single Protein Folding

Yongli Zhang, Junyi Jiao, Aleksander A. Rebane

Hidden Markov modeling (HMM) has revolutionized kinetic studies of macromolecules. However, results from HMM often violate detailed balance when applied to the transitions under thermodynamic equilibrium, and the consequence of such violation has not been well understood. Here, to our knowledge, we developed a new HMM method that satisfies detailed balance (HMM-DB) and optimizes model parameters by gradient search. We used free energy of stable and transition states as independent fitting parameters and considered both normal and skew normal distributions of the measurement noise. We validated our method by analyzing simulated extension trajectories that mimicked experimental data of single protein folding from optical tweezers. We then applied HMM-DB to elucidate kinetics of regulated SNARE zippering containing degenerate states. For both simulated and measured trajectories, we found that HMM-DB significantly reduced overfitting of short trajectories compared to the standard HMM based on an expectation-maximization algorithm, leading to more accurate and reliable model fitting by HMM-DB. We revealed how HMM-DB could be conveniently used to derive a simplified energy landscape of protein folding. Finally, we extended HMM-DB to correct the baseline drift in single-molecule trajectories. Together, we demonstrated an efficient, versatile, and reliable method of HMM for kinetics studies of macromolecules under thermodynamic equilibrium.


Cooperatively enhanced dipole forces from artificial atoms in trapped nanodiamonds

Mathieu L. Juan, Carlo Bradac, Benjamin Besga, Mattias Johnsson, Gavin Brennen, Gabriel Molina-Terriza & Thomas Volz
Optical trapping is a powerful tool to manipulate small particles, from micrometre-size beads in liquid environments1 to single atoms in vacuum2. The trapping mechanism relies on the interaction between a dipole and the electric field of laser light. In atom trapping, the dominant contribution to the associated force typically comes from the allowed optical transition closest to the laser wavelength, whereas for mesoscopic particles it is given by the polarizability of the bulk material. Here, we show that for nanoscale diamond crystals containing a large number of artificial atoms, nitrogen–vacancy colour centres, the contributions from both the nanodiamond and the colour centres to the optical trapping strength can be simultaneously observed in a noisy liquid environment. For wavelengths around the zero-phonon line transition of the colour centres, we observe a 10% increase of overall trapping strength. The magnitude of this effect suggests that due to the large density of centres, cooperative effects between the artificial atoms contribute to the observed modification of the trapping strength. Our approach may enable the study of cooperativity in nanoscale solid-state systems and the use of atomic physics techniques in the field of nano-manipulation.


Single-molecule studies of high-mobility group B architectural DNA bending proteins

Divakaran Murugesapillai, Micah J. McCauley, L. James MaherIII, Mark C. Williams
Protein–DNA interactions can be characterized and quantified using single molecule methods such as optical tweezers, magnetic tweezers, atomic force microscopy, and fluorescence imaging. In this review, we discuss studies that characterize the binding of high-mobility group B (HMGB) architectural proteins to single DNA molecules. We show how these studies are able to extract quantitative information regarding equilibrium binding as well as non-equilibrium binding kinetics. HMGB proteins play critical but poorly understood roles in cellular function. These roles vary from the maintenance of chromatin structure and facilitation of ribosomal RNA transcription (yeast high-mobility group 1 protein) to regulatory and packaging roles (human mitochondrial transcription factor A). We describe how these HMGB proteins bind, bend, bridge, loop and compact DNA to perform these functions. We also describe how single molecule experiments observe multiple rates for dissociation of HMGB proteins from DNA, while only one rate is observed in bulk experiments. The measured single-molecule kinetics reveals a local, microscopic mechanism by which HMGB proteins alter DNA flexibility, along with a second, much slower macroscopic rate that describes the complete dissociation of the protein from DNA.


Monday, November 21, 2016

Measurements and Predictions of Binary Component Aerosol Particle Viscosity

Young Chul Song, Allen E. Haddrell, Bryan R. Bzdek, Jonathan P. Reid, Thomas Bannan, David O. Topping, Carl Percival, and Chen Cai

Organic aerosol particles are known to often absorb/desorb water continuously with change in gas phase relative humidity (RH) without crystallization. Indeed, the prevalence of metastable ultraviscous liquid or amorphous phases in aerosol is well-established with solutes often far exceeding bulk phase solubility limits. Particles are expected to become increasingly viscous with drying, a consequence of the plasticizing effect of water. We report here measurements of the variation in aerosol particle viscosity with RH (equal to condensed phase water activity) for a range of organic solutes including alcohols (diols to hexols), saccharides (mono-, di-, and tri-), and carboxylic acids (di-, tri-, and mixtures). Particle viscosities are measured over a wide range (10–3 to 1010 Pa s) using aerosol optical tweezers, inferring the viscosity from the time scale for a composite particle to relax to a perfect sphere following the coalescence of two particles. Aerosol measurements compare well with bulk phase studies (well-within an order of magnitude deviation at worst) over ranges of water activity accessible to both. Predictions of pure component viscosity from group contribution approaches combined with either nonideal or ideal mixing reproduce the RH-dependent trends particularly well for the alcohol, di-, and tricarboxylic acid systems extending up to viscosities of 104 Pa s. By contrast, predictions overestimate the viscosity by many orders of magnitude for the mono-, di-, and trisaccharide systems, components for which the pure component subcooled melt viscosities are ≫1012 Pa s. When combined with a typical scheme for simulating the oxidation of α-pinene, a typical atmospheric pathway to secondary organic aerosol (SOA), these predictive tools suggest that the pure component viscosities are less than 106 Pa s for ∼97% of the 50,000 chemical products included in the scheme. These component viscosities are consistent with the conclusion that the viscosity of α-pinene SOA is most likely in the range 105 to 108 Pa s. Potential improvements to the group contribution predictive tools for pure component viscosities are considered.


The Power of Force: Insights into the Protein Folding Process Using Single-Molecule Force Spectroscopy

Jörg Schönfelder, David De Sancho, Raul Perez-Jimenez

One of the major challenges in modern biophysics is observing and understanding conformational changes during complex molecular processes, from the fundamental protein folding to the function of molecular machines. Single-molecule techniques have been one of the major driving forces of the huge progress attained in the last few years. Recent advances in resolution of the experimental setups, aided by theoretical developments and molecular dynamics simulations, have revealed a much higher degree of complexity inside these molecular processes than previously reported using traditional ensemble measurements. This review sums up the evolution of these developments and gives an outlook on prospective discoveries.


Generation of nanoscale anticounterfeiting patterns on silicon by optical trap assisted nanopatterning

T.-H. Chen, Y.-C. Tsai, R. Fardel and C. B. Arnold

Among the different strategies aimed at protecting products from counterfeiting, hidden security patterns are used by manufacturers to mark their products in a unique way. However, most anticounterfeiting patterns bear the risk of being reproduced by an unauthorized party who has gained knowledge of the exact technique and process parameters. In this paper, we use optical trap assisted nanopatterning to create unique security markings by taking advantage of statistical fluctuations when generating nanoscale features within the pattern. We image the patterns by optical microscopy, scanning electron microscopy, and atomic force microscopy and propose a three-level examination process that allows for an efficient yet highly secure authentication.


Optical force enhancement and annular trapping by plasmonic toroidal resonance in a double-disk metastructure

Ren-chao Jin, Jie Li, Ying-hua Wang, Ming-jie Zhu, Jia-qi Li, and Zheng-gao Dong

Optical forces can be enhanced by surface plasmon resonances with various interesting characteristics. Here, we numerically calculated the optical forces enhanced by a new kind of toroidal dipolar resonance in a double-disk metastructure. The results show that this kind of optical force is competitive with ordinary plasmonic forces and typically can reach−182.5𝑝𝑁𝜇𝑚2𝑚𝑊−1−182.5pNμm2mW−1. Influences of geometric parameters are discussed for the enhancement characteristic of optical force. Finally, we make a contrastive investigation on the optical trapping characteristic on a 5-nm-diameter nanoparticle, and show that the unique annular trapping region can be utilized for nanoscale applications.


Friday, November 18, 2016

Single Molecule Localization and Discrimination of DNA–Protein Complexes by Controlled Translocation Through Nanocapillaries

Roman D. Bulushev, Sanjin Marion, Ekaterina Petrova, Sebastian J. Davis, Sebastian J. Maerkl, and Aleksandra Radenovic

Through the use of optical tweezers we performed controlled translocations of DNA–protein complexes through nanocapillaries. We used RNA polymerase (RNAP) with two binding sites on a 7.2 kbp DNA fragment and a dCas9 protein tailored to have five binding sites on λ-DNA (48.5 kbp). Measured localization of binding sites showed a shift from the expected positions on the DNA that we explained using both analytical fitting and a stochastic model. From the measured force versus stage curves we extracted the nonequilibrium work done during the translocation of a DNA–protein complex and used it to obtain an estimate of the effective charge of the complex. In combination with conductivity measurements, we provided a proof of concept for discrimination between different DNA–protein complexes simultaneous to the localization of their binding sites.


Precise Sorting of Gold Nanoparticles in a Flowing System

Wei Wu, Xiaoqiang Zhu, Yunfeng Zuo, Li Liang, Shunping Zhang, Xuming Zhang, and Yi Yang

Precise sorting of gold nanoparticles is important, but it still remains a big challenge. Traditional methods such as centrifugation can separate nanoparticles with a high throughput but at the cost of low precision. Optical tweezers enable the precise manipulation of a single nanoparticle in steady liquid environments. However, this method may become problematic when dealing with a considerable amount of nanoparticles in a flowing system due to the difficulties in balancing the additional Stokes forces by the fast velocity of streams and in controlling all dispersed nanoparticles with disorderly positions. Here, we exploit optical and hydrodynamic forces to sort gold nanoparticles in the flowing system, obtaining simultaneously high precision and considerable throughput. This is accomplished by utilizing opposite impinging streams to generate a stagnation point, near which the flow velocity becomes very small to reduce the Stokes force and to prolong the optical acting time. Nanoparticles of different sizes, confined in a narrow region by the hydrodynamic focusing, can then be separated by a laser beam of moderate power. Experimental demonstrations have been presented by sorting gold nanoparticles with diameters of 50 nm from those of 100 nm, and 100 nm from 200 nm. The sorting fidelities is ≥92% for the 50/100 nm combination and ≥86% for the 100/200 nm set, with a sorting throughput of 300 particles/min. Sorting of gold nanoparticles with smaller heterogeneity (50 and 70 nm) has also been realized with a lower throughput of <100 particles/min. Our method can also be extended to separate nanoparticles of different shapes and compositions, which shows its great promise in the fields of plasmonics and nanophotonics.


Lorentz force and the optical pulling of multiple rayleigh particles outside the dielectric cylindrical waveguides

M.R.C. Mahdy, M. Q. Mehmood, Weiqiang Ding, Tianhang Zhang, Zhi Ning Chen

The stimulating connection between the counter-intuitive optical pulling effects and the Lorentz force has not been investigated in literature. This work demonstrates that multiple absorbing or non-absorbing dielectric Rayleigh objects can be pulled locally with gradientless travelling waves outside a finite-sized cylindrical nano or micro waveguide, if it is made up of a hollow core along with the cladding of at least two different dielectrics of appropriate refractive indices. Lorentz force analysis reveals that the bound surface charges of Rayleigh scatterer experience backward force, which overcomes the positive bulk force and ultimately results in the net pulling of the scatterer for several spatial regions outside the waveguide. Finally, in order to control the pulling of multiple Rayleigh particles based on scattering force and binding force, we have proposed a possible cylindrical coupler set-up. This work may open a new window of optical pulling force due to the exclusion of conventional structured tractor beams along with the artificial exotic matters.


Holographic tracking and sizing of optically trapped microprobes in diamond anvil cells

F. Saglimbeni, S. Bianchi, G. Gibson, R. Bowman, M. Padgett, and R. Di Leonardo

We demonstrate that Digital Holographic Microscopy can be used for accurate 3D tracking and sizing of a colloidal probe trapped in a diamond anvil cell (DAC). Polystyrene beads were optically trapped in water up to Gigapascal pressures while simultaneously recording in-line holograms at 1 KHz frame rate. Using Lorenz-Mie scattering theory to fit interference patterns, we detected a 10% shrinking in the bead’s radius due to the high applied pressure. Accurate bead sizing is crucial for obtaining reliable viscosity measurements and provides a convenient optical tool for the determination of the bulk modulus of probe material. Our technique may provide a new method for pressure measurements inside a DAC.


Thursday, November 17, 2016

Orbital-angular-momentum transfer to optically levitated microparticles in vacuum

Michael Mazilu, Yoshihiko Arita, Tom Vettenburg, Juan M. Auñón, Ewan M. Wright, and Kishan Dholakia

We demonstrate the transfer of orbital angular momentum to an optically levitated microparticle in vacuum. The microparticle is placed within a Laguerre-Gaussian beam and orbits the annular beam profile with increasing angular velocity as the air drag coefficient is reduced. We explore the particle dynamics as a function of the topological charge of the levitating beam. Our results reveal that there is a fundamental limit to the orbital angular momentum that may be transferred to a trapped particle, dependent upon the beam parameters and inertial forces present.


Unraveling the physical chemistry and the mixed binding modes of complex DNA ligands by single molecule stretching experiments

W. F. P. Bernal, E. F. Silva and M. S. Rocha

In this work we present a complete methodology to unravel the physical chemistry and the mixed binding modes of complex DNA ligands. Single molecule stretching experiments were performed with complexes formed between a DNA binding drug that exhibits multiple mixed binding modes (Berenil) and the biopolymer. From these experiments we determine the changes of the two basic mechanical properties, the contour and persistence lengths, as a function of the drug concentration in the sample. Combining a modeling analysis for the two mechanical properties, we were able to extract the physicochemical parameters of the interaction and to determine the effective binding mechanisms. In particular, we have shown that in this case the binding modes can be modulated by changing the ionic strength of the surrounding buffer: for high ionic strengths (150 mM), Berenil behaves as a typical minor groove ligand in its interaction with λ-DNA; while, for low ionic strengths (10 mM), the drug also partially intercalates into the double-helix. The methodology developed in the present analysis can be promptly applied to other complex DNA ligands, therefore allowing one to investigate and decouple different binding mechanisms.


DNA as a Model for Probing Polymer Entanglements: Circular Polymers and Non-Classical Dynamics

Kathryn Regan, Shea Ricketts and Rae M. Robertson-Anderson

Double-stranded DNA offers a robust platform for investigating fundamental questions regarding the dynamics of entangled polymer solutions. The exceptional monodispersity and multiple naturally occurring topologies of DNA, as well as a wide range of tunable lengths and concentrations that encompass the entanglement regime, enable direct testing of molecular-level entanglement theories and corresponding scaling laws. DNA is also amenable to a wide range of techniques from passive to nonlinear measurements and from single-molecule to bulk macroscopic experiments. Over the past two decades, researchers have developed methods to directly visualize and manipulate single entangled DNA molecules in steady-state and stressed conditions using fluorescence microscopy, particle tracking and optical tweezers. Developments in microfluidics, microrheology and bulk rheology have also enabled characterization of the viscoelastic response of entangled DNA from molecular levels to macroscopic scales and over timescales that span from linear to nonlinear regimes. Experiments using DNA have uniquely elucidated the debated entanglement properties of circular polymers and blends of linear and circular polymers. Experiments have also revealed important lengthscale and timescale dependent entanglement dynamics not predicted by classical tube models, both validating and refuting new proposed extensions and alternatives to tube theory and motivating further theoretical work to describe the rich dynamics exhibited in entangled polymer systems.


H2A.Z controls the stability and mobility of nucleosomes to regulate expression of the LH genes

Sergei Rudnizky, Adaiah Bavly, Omri Malik, Lilach Pnueli, Philippa Melamed & Ariel Kaplan

The structure and dynamics of promoter chromatin have a profound effect on the expression levels of genes. Yet, the contribution of DNA sequence, histone post-translational modifications, histone variant usage and other factors in shaping the architecture of chromatin, and the mechanisms by which this architecture modulates expression of specific genes are not yet completely understood. Here we use optical tweezers to study the roles that DNA sequence and the histone variant H2A.Z have in shaping the chromatin landscape at the promoters of two model genes, Cga and Lhb. Guided by MNase mapping of the promoters of these genes, we reconstitute nucleosomes that mimic those located near the transcriptional start site and immediately downstream (+1), and measure the forces required to disrupt these nucleosomes, and their mobility along the DNA sequence. Our results indicate that these genes are basally regulated by two distinct strategies, making use of H2A.Z to modulate separate phases of transcription, and highlight how DNA sequence, alternative histone variants and remodelling machinery act synergistically to modulate gene expression.


Ultralow-Power Electronic Trapping of Nanoparticles with Sub-10 nm Gold Nanogap Electrodes

Avijit Barik, Xiaoshu Chen, and Sang-Hyun Oh

We demonstrate nanogap electrodes for rapid, parallel, and ultralow-power trapping of nanoparticles. Our device pushes the limit of dielectrophoresis by shrinking the separation between gold electrodes to sub-10 nm, thereby creating strong trapping forces at biases as low as the 100 mV ranges. Using high-throughput atomic layer lithography, we manufacture sub-10 nm gaps between 0.8 mm long gold electrodes and pattern them into individually addressable parallel electronic traps. Unlike pointlike junctions made by electron-beam lithography or larger micron-gap electrodes that are used for conventional dielectrophoresis, our sub-10 nm gold nanogap electrodes provide strong trapping forces over a mm-scale trapping zone. Importantly, our technology solves the key challenges associated with traditional dielectrophoresis experiments, such as high voltages that cause heat generation, bubble formation, and unwanted electrochemical reactions. The strongly enhanced fields around the nanogap induce particle-transport speed exceeding 10 μm/s and enable the trapping of 30 nm polystyrene nanoparticles using an ultralow bias of 200 mV. We also demonstrate rapid electronic trapping of quantum dots and nanodiamond particles on arrays of parallel traps. Our sub-10 nm gold nanogap electrodes can be combined with plasmonic sensors or nanophotonic circuitry, and their low-power electronic operation can potentially enable high-density integration on a chip as well as portable biosensing.


Thursday, November 10, 2016

Fluctuations of a membrane nanotube revealed by high-resolution force measurements

F. Valentino, P. Sens, J. Lemière, A. Allard, T. Betz, C. Campillo and C. Sykes

Pulling membrane nanotubes from liposomes presents a powerful method to gain access to membrane mechanics. Here we extend classical optical tweezers studies to infer membrane nanotube dynamics with high spatial and temporal resolution. We first validate our force measurement setup by accurately measuring the bending modulus of EPC membrane in tube pulling experiments. Then we record the position signal of a trapped bead when it is connected, or not, to a tube. We derive the fluctuation spectrum of these signals and find that the presence of a membrane nanotube induces higher fluctuations, especially at low frequencies (10–1000 Hz). We analyse these spectra by taking into account the peristaltic modes of nanotube fluctuations. This analysis provides a new experimental framework for a quantitative study of the fluctuations of nanotubular membrane structures that are present in living cells, and now classically used for in vitro biomimetic approaches.


HIF Stabilization Weakens Primary Cilia

Andrew Resnick

Although solitary or sensory cilia are present in most cells of the body and their existence has been known since the sixties, very little is known about their functions. One suspected function is fluid flow sensing- physical bending of cilia produces an influx of Ca++, which can then result in a variety of activated signaling pathways. Defective cilia and ciliary-associated proteins have been shown to result in cystic diseases. Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a progressive disease, typically appearing in the 5th decade of life and is one of the most common monogenetic inherited human diseases, affecting approximately 600,000 people in the United States. Because the mechanical properties of cilia impact their response to applied flow, we asked how the stiffness of cilia can be controlled pharmacologically. We performed an experiment subjecting cilia to Taxol (a microtubule stabilizer) and CoCl2 (a HIF stabilizer to model hypoxia). Madin-Darby Canine Kidney (MDCK) cells were selected as our model system. After incubation with a selected pharmacological agent, cilia were optically trapped and the bending modulus measured. We found that HIF stabilization significantly weakens cilia. These results illustrate a method to alter the mechanical properties of primary cilia and potentially alter the flow sensing properties of cilia.


DNA looping mediates nucleosome transfer

Lucy D. Brennan, Robert A. Forties, Smita S. Patel & Michelle D. Wang

Proper cell function requires preservation of the spatial organization of chromatin modifications. Maintenance of this epigenetic landscape necessitates the transfer of parental nucleosomes to newly replicated DNA, a process that is stringently regulated and intrinsically linked to replication fork dynamics. This creates a formidable setting from which to isolate the central mechanism of transfer. Here we utilized a minimal experimental system to track the fate of a single nucleosome following its displacement, and examined whether DNA mechanics itself, in the absence of any chaperones or assembly factors, may serve as a platform for the transfer process. We found that the nucleosome is passively transferred to available dsDNA as predicted by a simple physical model of DNA loop formation. These results demonstrate a fundamental role for DNA mechanics in mediating nucleosome transfer and preserving epigenetic integrity during replication.


Atom-by-atom assembly of defect-free one-dimensional cold atom arrays

Manuel Endres, Hannes Bernien, Alexander Keesling, Harry Levine, Eric R. Anschuetz, Alexandre Krajenbrink, Crystal Senko, Vladan Vuletic, Markus Greiner, Mikhail D. Lukin

The realization of large-scale fully controllable quantum systems is an exciting frontier in modern physical science. We use atom-by-atom assembly to implement a platform for the deterministic preparation of regular one-dimensional arrays of individually controlled cold atoms. In our approach, a measurement and feedback procedure eliminates the entropy associated with probabilistic trap occupation and results in defect-free arrays of over 50 atoms in less than 400 milliseconds. The technique is based on fast, real-time control of 100 optical tweezers, which we use to arrange atoms in desired geometric patterns and to maintain these configurations by replacing lost atoms with surplus atoms from a reservoir. This bottom-up approach may enable controlled engineering of scalable many-body systems for quantum information processing, quantum simulations, and precision measurements.


The evolution of multivalent nanoparticle adhesion via specific molecular interactions

Mingqiu Wang, Shreyas R. Ravindranath, Maha K Rahim, Elliot Botvinick, and Jered Brackston Haun
The targeted delivery of nanoparticle carriers holds tremendous potential to transform the detection and treatment of diseases. A major attribute of nanoparticles is the ability to form multiple bonds with target cells, which greatly improves adhesion strength. However, multivalent binding of nanoparticles is still poorly understood, particularly from a dynamic perspective. In previous experimental work, we studied the kinetics of nanoparticle adhesion and found that the rate of detachment decreased over time. Here, we have applied the Adhesive Dynamics simulation framework to investigate binding dynamics between an antibody-conjugated, 200 nm diameter sphere and ICAM-1 on a surface at the scale of individual bonds. We found that Nano Adhesive Dynamics (NAD) simulations could replicate the time-varying nanoparticle detachment behavior that we observed in experiments. As expected, this was correlated with a steady increase in mean bond number with time, but this was only attributed to bond accumulation during the first second that nanoparticles were bound. Longer-term increases in bond number instead manifested from nanoparticle detachment serving as a selection mechanism to eliminate nanoparticles that had randomly been confined to lower bond valencies. Thus, time-dependent nanoparticle detachment reflects an evolution of the remaining nanoparticle population towards higher overall bond valency. We also found that NAD simulations precisely matched experiments whenever mechanical force loads on bonds was high enough to directly induce rupture. These mechanical forces were in excess of 300 pN and primarily arose from the Brownian motion of the nanoparticle, but we also identified a valency-dependent contribution from bonds pulling on each other. In summary, we have achieved excellent kinetic consistency between NAD simulations and experiments, which has revealed new insights into the dynamics and biophysics of multivalent nanoparticle adhesion. In future work we will seek to leverage the simulation as a design tool for optimizing targeted nanoparticle agents.


Wednesday, November 9, 2016

Single-molecule measurements of viral ssRNA packaging

Kalle J. Hanhijärvi, Gabija Ziedaite, Dennis H. Bamford, Edward Hæggström and Minna M. Poranen

Genome packaging of double-stranded RNA (dsRNA) phages has been widely studied using biochemical and molecular biology methods. We adapted the existing in vitro packaging system of one such phage for single-molecule experimentation. To our knowledge, this is the first attempt to study the details of viral RNA packaging using optical tweezers. Pseudomonas phage phi6 is a dsRNA virus with a tripartite genome. Positive-sense (+) single-stranded RNA (ssRNA) genome precursors are packaged into a preformed procapsid (PC), where negative-strands are synthesized. We present single-molecule measurements of the viral ssRNA packaging by the phi6 PC. Our data show that packaging proceeds intermittently in slow and fast phases, which likely reflects differences in the unfolding of the RNA secondary structures of the ssRNA being packaged. Although the mean packaging velocity was relatively low (0.07–0.54 nm / s), packaging could reach 4.62 nm / s during the fast packaging phase.


In situ single-atom array synthesis using dynamic holographic optical tweezers

Hyosub Kim, Woojun Lee, Han-gyeol Lee, Hanlae Jo, Yunheung Song & Jaewook Ahn

Establishing a reliable method to form scalable neutral-atom platforms is an essential cornerstone for quantum computation, quantum simulation and quantum many-body physics. Here we demonstrate a real-time transport of single atoms using holographic microtraps controlled by a liquid-crystal spatial light modulator. For this, an analytical design approach to flicker-free microtrap movement is devised and cold rubidium atoms are simultaneously rearranged with 2N motional degrees of freedom, representing unprecedented space controllability. We also accomplish an in situ feedback control for single-atom rearrangements with the high success rate of 99% for up to 10 μm translation. We hope this proof-of-principle demonstration of high-fidelity atom-array preparations will be useful for deterministic loading of N single atoms, especially on arbitrary lattice locations, and also for real-time qubit shuttling in high-dimensional quantum computing architectures.


Cell-structure specific necrosis by optical-trap induced intracellular nuclear oscillation

X.X. Sun, Z.L. Zhou, C.H. Man, A.Y.H. Leung, A.H.W. Ngan

A drug-free procedure for killing malignant cells in a cell-type specific manner would represent a significant breakthrough for leukemia treatment. Here, we show that mechanically vibrating a cell in a specific oscillation condition can significantly promote necrosis. Specifically, oscillating the cell by a low-power laser trap at specific frequencies of a few Hz was found to result in increased death rate of 50% or above in different types of myelogenous leukemia cells, while normal leukocytes showed very little response to similar laser manipulations. The alteration of cell membrane permeability and cell volume, detected from ethidium bromide staining and measurement of intracellular sodium ion concentration, together with the observed membrane blebbing within 10 minutes, suggest cell necrosis. Mechanics modelling reveals severe distortion of the cytoskeleton cortex at frequencies in the same range for peaked cell death. The disruption of cell membrane leading to cell death is therefore due to the cortex distortion, and the frequency at which this becomes significant is cell-type specific. Our findings lay down a new concept for treating leukemia based on vibration induced disruption of membrane in targeted malignant cells.


Guided transport of nanoparticles by plasmonic nanowires

Cui Yang, Pan Deng, Lianming Tong and Hongxing Xu

Here we report the optical trapping and directional transport of nanoparticles in aqueous by plasmonic nanowires. The laser illuminated on one-end of a silver nanowire and excited the localized and propagating surface plasmons. Optical forces were induced by the surface plasmons and can trap nanoparticles in aqueous. Interestingly, the trapped nanoparticles moved along the silver nanowires from the trapping site to the excitation spot of the laser. Such movements of nanoparticles were also observed on curved nanowires, on which the trajectories of particles were explicitly determined by the shape of nanowires. More importantly, for a V-shaped silver nanowire, the direction of the movement can be modulated by the polarization of the incident laser. The direction of the movement was opposite to the prediction by the scattering force due to the propagation of surface plasmons, and the driving force could involve the thermal convection of local fluid due to the heating effect. Our findings indicate a novel approach to transporting nanoparticles by plasmomic waveguides in aqueous.


Tuesday, November 8, 2016

Chiral particles in the dual-beam optical trap

Oto Brzobohatý, Raúl Josué Hernández, Stephen Simpson, Alfredo Mazzulla, Gabriella Cipparrone, and Pavel Zemánek
We investigate the dynamics of chiral microparticles in a dual-beam optical trap. The chiral particles have the structure of spherical chiral microresonators, with a reflectance deriving from the supramolecular helicoidal arrangement. Due to the strong asymmetric response of the particles to light with a specific helicity and wavelength, their trapping position and rotational frequency can be controlled by proper combination of the polarization state of the two light beams. Here symmetric and asymmetric polarization configurations of dual- interfering beam traps have been investigated. Based on the polarization controlled asymmetric transmission of the chiral particles, a tunable wash-board potential is created enabling the control of the trapping position along the beams axis. Asymmetric configurations display polarization controlled rotation of the trapped particles. Optical binding of rotating particles exhibits a complex dynamics.


Precise control and measurement of solid–liquid interfacial temperature and viscosity using dual-beam femtosecond optical tweezers in the condensed phase

Dipankar Mondal, Paresh Mathur and Debabrata Goswami

We present a novel method of microrheology based on femtosecond optical tweezers, which in turn enables us to directly measure and control in situ temperature at microscale volumes at the solid–liquid interface. A noninvasive pulsed 780 nm trapped bead spontaneously responds to changes in its environment induced by a co-propagating 1560 nm pulsed laser due to mutual energy transfer between the solvent molecules and the trapped bead. Strong absorption of the hydroxyl group by the 1560 nm laser creates local heating in individual and binary mixtures of water and alcohols. “Hot Brownian motion” of the trapped polystyrene bead is reflected in the corner frequency deduced from the power spectrum. Changes in corner frequency values enable us to calculate the viscosity as well as temperature at the solid–liquid interface. We show that these experimental results can also be theoretically ratified.


Microtrap on a concave grating reflector for atom trapping

Hui Zhang, Tao Li, Ya-Ling Yin, Xing-Jia Li, Yong Xia and Jian-Ping Yin

We propose a novel scheme of optical confinement for atoms by using a concave grating reflector. The two-dimension grating structure with a concave surface shape exhibits strong focusing ability under radially polarized illumination. Especially, the light intensity at the focal point is about 100 times higher than that of the incident light. Such a focusing optical field reflected from the curved grating structure can provide a deep potential to trap cold atoms. We discuss the feasibility of the structure serving as an optical dipole trap. Our results are as follows. (i) Van der Waals attraction potential to the surface of the structure has a low effect on trapped atoms. (ii) The maximum trapping potential is ~ 1.14 mK in the optical trap, which is high enough to trap cold 87Rb atoms from a standard magneto-optical trap with a temperature of 120 μK, and the maximum photon scattering rate is lower than 1/s. (iii) Such a microtrap array can also manipulate and control cold molecules, or microscopic particles.


Laser-Induced Motion of a Nanofluid in a Micro-Channel

Tran X. Phuoc, Mehrdad Massoudi and Ping Wang
Since a photon carries both energy and momentum, when it interacts with a particle, photon-particle energy and momentum transfer occur, resulting in mechanical forces acting on the particle. In this paper we report our theoretical study on the use of a laser beam to manipulate and control the flow of nanofluids in a micro-channel. We calculate the velocity induced by a laser beam for TiO2, Fe2O3, Al2O3 MgO, and SiO2 nanoparticles with water as the base fluid. The particle diameter is 50 nm and the laser beam is a 4 W continuous beam of 6 mm diameter and 532 nm wavelength. The results indicate that, as the particle moves, a significant volume of the surrounding water (up to about 8 particle diameters away from the particle surface) is disturbed and dragged along with the moving particle. The results also show the effect of the particle refractive index on the particle velocity and the induced volume flow rate. The velocity and the volume flowrate induced by the TiO2 nanoparticle (refractive index n = 2.82) are about 0.552 mm/s and 9.86 fL, respectively, while those induced by SiO2 (n = 1.46) are only about 7.569 μm/s and 0.135, respectively.


Monday, November 7, 2016

Alternative modes of client binding enable functional plasticity of Hsp70

Alireza Mashaghi, Sergey Bezrukavnikov, David P. Minde, Anne S. Wentink, Roman Kityk, Beate Zachmann-Brand, Matthias P. Mayer, Günter Kramer, Bernd Bukau & Sander J. Tans

The Hsp70 system is a central hub of chaperone activity in all domains of life. Hsp70 performs a plethora of tasks, including folding assistance, protection against aggregation, protein trafficking, and enzyme activity regulation1, 2, 3, 4, 5, and interacts with non-folded chains, as well as near-native, misfolded, and aggregated proteins6, 7, 8, 9, 10. Hsp70 is thought to achieve its many physiological roles by binding peptide segments that extend from these different protein conformers within a groove that can be covered by an ATP-driven helical lid11, 12, 13, 14, 15. However, it has been difficult to test directly how Hsp70 interacts with protein substrates in different stages of folding and how it affects their structure. Moreover, recent indications of diverse lid conformations in Hsp70–substrate complexes raise the possibility of additional interaction mechanisms15, 16, 17, 18. Addressing these issues is technically challenging, given the conformational dynamics of both chaperone and client, the transient nature of their interaction, and the involvement of co-chaperones and the ATP hydrolysis cycle19. Here, using optical tweezers, we show that the bacterial Hsp70 homologue (DnaK) binds and stabilizes not only extended peptide segments, but also partially folded and near-native protein structures. The Hsp70 lid and groove act synergistically when stabilizing folded structures: stabilization is abolished when the lid is truncated and less efficient when the groove is mutated. The diversity of binding modes has important consequences: Hsp70 can both stabilize and destabilize folded structures, in a nucleotide-regulated manner; like Hsp90 and GroEL, Hsp70 can affect the late stages of protein folding; and Hsp70 can suppress aggregation by protecting partially folded structures as well as unfolded protein chains. Overall, these findings in the DnaK system indicate an extension of the Hsp70 canonical model that potentially affects a wide range of physiological roles of the Hsp70 system.


Dynamic viscosity mapping of the oxidation of squalene aerosol particles

Athanasios Athanasiadis, Clare Fitzgerald, Nicholas M. Davidson, Chiara Giorio, Stanley W. Botchway, Andrew D. Ward, Markus Kalberer, Francis D. Pope and Marina K. Kuimova

Organic aerosols (OAs) play important roles in multiple atmospheric processes, including climate change, and can impact human health. The physico-chemical properties of OAs are important for all these processes and can evolve through reactions with various atmospheric components, including oxidants. The dynamic nature of these reactions makes it challenging to obtain a true representation of their composition and surface chemistry. Here we investigate the microscopic viscosity of the model OA composed of squalene, undergoing chemical aging. We employ Fluorescent Lifetime Imaging Microscopy (FLIM) in conjunction with viscosity sensitive probes termed molecular rotors, in order to image the changes in microviscosity in real time during oxidation with ozone and hydroxyl radicals, which are two key oxidising species in the troposphere. We also recorded the Raman spectra of the levitated particles to follow the reactivity during particle ozonolysis. The levitation of droplets was achieved via optical trapping that enabled simultaneous levitation and measurement via FLIM or Raman spectroscopy and allowed the true aerosol phase to be probed. Our data revealed a very significant increase in viscosity of the levitated squalene droplets upon ozonolysis, following their transformation from the liquid to solid phase that was not observable when the oxidation was carried out on coverslip mounted droplets. FLIM imaging with sub-micron spatial resolution also revealed spatial heterogeneity in the viscosity distribution of oxidised droplets. Overall, a combination of molecular rotors, FLIM and optical trapping is able to provide powerful insights into OA chemistry and the microscopic structure that enables the dynamic monitoring of microscopic viscosity in aerosol particles in their true phase.


Single-Molecule Optical-Trapping Techniques to Study Molecular Mechanisms of a Replisome

B. Sun, M.D. Wang

The replisome is a multiprotein molecular machinery responsible for the replication of DNA. It is composed of several specialized proteins each with dedicated enzymatic activities, and in particular, helicase unwinds double-stranded DNA and DNA polymerase catalyzes the synthesis of DNA. Understanding how a replisome functions in the process of DNA replication requires methods to dissect the mechanisms of individual proteins and of multiproteins acting in concert. Single-molecule optical-trapping techniques have proved to be a powerful approach, offering the unique ability to observe and manipulate biomolecules at the single-molecule level and providing insights into the mechanisms of molecular motors and their interactions and coordination in a complex. Here, we describe a practical guide to applying these techniques to study the dynamics of individual proteins in the bacteriophage T7 replisome, as well as the coordination among them. We also summarize major findings from these studies, including nucleotide-specific helicase slippage and new lesion bypass pathway in T7 replication.


Direct Visualization of Helicase Dynamics Using Fluorescence Localization and Optical Trapping

C.-T. Lin, T. Ha

Helicases control the accessibility of single-stranded (ss) nucleic acid (NA) generated as a transient intermediate during almost every step in cells related to nucleic acid metabolisms. For subsequent processing, however, helicases need to adjust the pace of unwinding adequately to avoid ssNA exposure to nucleases. Therefore, understanding how the unwinding process of helicases is regulated is crucial to address genome integrity and repair mechanisms. Using single-molecule fluorescence-force spectroscopy with fluorescence localization, we recently observed the stoichiometry of UvrD helicase, which determines the functions of UvrD: translocation and unwinding. For the first time, we provide direct evidence that a UvrD dimer is required to initiate the unwinding pathway. Moreover, with subpixel precision of fluorescence localization, the dynamic parameters of helicases can be obtained directly. Here, we present detailed single-molecule assays for observing the biochemical activities of helicases in real time and revealing how mechanical forces are involved in protein–nucleic acid interactions. These single-molecule approaches are generally applicable to many other protein–nucleic acid systems.


Numerical analysis of an optical nanoscale particles trapping device based on a slotted nanobeam cavity

Senlin Zhang, Zhengdong Yong, Yaocheng Shi & Sailing He

A slotted nanobeam cavity (SNC) is utilized to trap a polystyrene (PS) particle with a radius of only 2 nm. The carefully designed SNC shows an ultrahigh Q factor of 4.5 × 107 while maintaining a small mode volume of 0.067(λ/nwater)3. Strongly enhanced optical trapping force is numerically demonstrated when the 2 nm PS particle is introduced into the central, slotted part of the SNC. In the vertical direction, the numerical calculation results show that a trapping stiffness of 0.4 pN/(nm · mW) around the equilibrium position and a trapping potential barrier of ~2000 kBT/mW can be reached. To our best knowledge, the trapping capability (trapping stiffness and trapping potential barrier) of the proposed structure significantly outperforms the theoretical results of those in previously reported work. In addition, the SNC system does not suffer from the metal induced heat issue that restricts the performance of state-of-the-art optical trapping systems involving plasmonic enhancement. Based on the proposed cavity, applications such as lab-on-a-chip platforms for nanoscale particle trapping and analysis can be expected in future.


Tuesday, November 1, 2016

Characterization of drug effect on leukemia cells through single cell assay with optical tweezers and dielectrophoresis

Jundi Hou ; Tao Luo ; Ka Lam Ng ; Raymond Liang ; Anskar Y. H. Leung ; Dong Sun

One of the greatest challenges in acute myeloid leukemia (AML) treatment is preventing relapse. Leukemia cells can hide in bone marrow niche or vascular niche. Hence, many chemical drugs cannot kill these cells. To characterize migration and adhesion properties of leukemia cells in specific niches, CXCR4/SDF-1α signal pathway has been widely used for investigation. AMD3100 is treated as one of the most common chemical drugs that can inhibit this signal. In the current study, we particularly investigate the effect of AMD3100 on the adhesion property of leukemia cells on stromal cells by using engineering tools, namely, optical tweezers (OT) and dielectrophoresis (DEP), to probe single cell property. AMD3100 not only inhibits the CXCR4/SDF-1α signal pathway but also reduces gene expression of CXCR4 and VLA4 on leukemia cells. The drug also softens leukemia cells. This work provides a new way to investigate cell behavior under drug treatment. The use of combined engineering tools will benefit drug discovery and assessment for leukemia treatment.


Cooperative Optical Trapping and Manipulation of Multiple Cells With Robot Tweezers

Xiang Li; Chien Chern Cheah; Quang Minh Ta

While several control schemes and automation techniques have been proposed for coordination and manipulation of multiple cells using optical tweezers, open-loop control methods are always utilized by treating the positions of lasers as the control inputs instead of feedback variables. As the positions of laser beams are not utilized in feedback control, it is, therefore, assumed in the literature that the multiple cells are always trapped by the laser beams throughout the manipulation task. However, the control techniques fail when the cells are not initially trapped by the laser beams, or when the laser beams move too fast, such that the cells escape from the traps during manipulation. This paper presents a closed-loop control formulation and strategy for optical manipulation of multiple cells, based on the cooperative movement of the motorized stage and the beam steering system. The closed-loop control method enables the trapping operation to be automatically activated whenever the cells are not inside the optical traps. The proposed controller does not require exact knowledge on the dynamic parameters and the varying trapping stiffness. Lyapunov methods are employed in the stability analysis of the optical tweezers system. Experimental results are presented to illustrate the performance of the proposed controller.


Potentialities of laser trapping and manipulation of blood cells in hemorheologic research

Priezzhev, A.; Lee, K.

Laser trapping and manipulation of blood cells without mechanical contact have become feasible with implication of laser tweezers. They open up new horizons for the hemorheologic researches, offer new possibilities for studying live cells interactions on individual cell level under the influence of different endogenous and exogenous factors. The operation principle of laser tweezers is based on the property of strongly focused laser beam to act on a dielectric microparticle located in the vicinity of the beam waist with a force that drives the particle to the equilibrium location and holds it there. If the beam waist position is manipulated, so is the position of the particle. The displacement of the particle from the equilibrium position by external forces can be calibrated so that these forces can be precisely measured in the range ca. 0.1–100 pN. This is the range of forces of elastic deformation of blood cells and of their interaction with each other and with vessel walls. Being able to measure these forces without mechanical contact allows for studying on single cell level the mechanisms of interactions that was impossible earlier. Here we discuss the basic features of these techniques and give some examples of challenging hemorheologic studies.


Nucleosomal arrangement affects single-molecule transcription dynamics

Veronika Fitz, Jaeoh Shin, Christoph Ehrlich, Lucas Farnung, Patrick Cramer, Vasily Zaburdaev, and Stephan W. Grill
In eukaryotes, gene expression depends on chromatin organization. However, how chromatin affects the transcription dynamics of individual RNA polymerases has remained elusive. Here, we use dual trap optical tweezers to study single yeast RNA polymerase II (Pol II) molecules transcribing along a DNA template with two nucleosomes. The slowdown and the changes in pausing behavior within the nucleosomal region allow us to determine a drift coefficient, χ, which characterizes the ability of the enzyme to recover from a nucleosomal backtrack. Notably, χ can be used to predict the probability to pass the first nucleosome. Importantly, the presence of a second nucleosome changes χ in a manner that depends on the spacing between the two nucleosomes, as well as on their rotational arrangement on the helical DNA molecule. Our results indicate that the ability of Pol II to pass the first nucleosome is increased when the next nucleosome is turned away from the first one to face the opposite side of the DNA template. These findings help to rationalize how chromatin arrangement affects Pol II transcription dynamics.